Use of photosynthetic pigment stabilizing agents to regulate ripening and quality in fruits and vegetables

ABSTRACT

Methods and compositions to temporarily delay the ripening, maturation, senescence and pathogen infection of produce such as fruits, vegetables and ornamental plants are provided. The methods involve the application of a photosynthetic pigment and/or membrane stabilizing chemical (e.g. glycine betaine) to produce which is identified as in need of such delay. Application may be prior to harvest, at or during harvest or post-harvest.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims benefit of U.S. provisional patent application61/489,103, filed May 23, 2011, and claims benefit of and is acontinuation-in-part of International patent application PCT/US12/39038,filed May 23, 2012, the complete contents of both of which are herebyincorporated by reference.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The invention generally relates to methods for delaying the abscission,preharvest drop, ripening, maturation, senescence or pathogen entry ofproduce such as fruit and vegetables. In particular, the inventionprovides a method of temporarily preventing or delaying the ripening ormaturation of produce by the pre- or post-harvest application of aphotosynthetic pigment or membrane stabilizing agent and osmoregulaorsuch as glycine betaine.

2. Background of the Invention

Postharvest treatment with the synthetic molecule 1-methylcyclopropene(1-MCP) is used by the fruit industry to slow the ripening process offruit during storage. 1-MCP treatment extends the shelf life of treatedfruit, thereby increasing the availability of fresh fruit to consumersthroughout much of the year. 1-MCP is structurally related to thenatural plant hormone ethylene and slows ripening by binding to ethylenereceptors, thereby blocking the binding of ethylene and preventing aripening response to the hormone. In some cases, over time, the effectof 1-MCP treatment wanes and ripening of the treated commodity canoccur. The use of 1-MCP in this manner is described in U.S. Pat. No.6,017,849, the complete contents of which are hereby incorporated byreference in entirety.

Unfortunately, certain fruits do not respond well to 1-MCP. For example,applications of 1-MCP have had mixed success in pear storage, oftenyielding pears that are unable to ripen at all after treatment. Theresulting pears remain hard and have relatively low levels of solublesugars, which impair their taste and appeal to consumers. For these andother types of fruit, there is a need in the industry to providealternative methods for the treatment of fruit to slow the ripeningprocess. It would also be desirable to have available an anti-ripeningagent that is a natural product rather than a synthetic such as 1-MCP.

Several attempts to control or delay ripening of fruits and vegetablesduring transport and storage have been made. For example, U.S. Pat. Nos.6,124,525, 6,080,914, 5,952,546 and 5,512,466 describe various geneticmodifications of plants that may be used to alter the ripening process.However, the use of genetically modified organisms is generally notpopular with the public at large, may have far-reaching andunpredictable environmental consequences, and requires a high degree ofgovernment regulation.

U.S. Pat. No. 7,943,549 to Pierce et al. describes methods for delayinga plant development process by exposing a plant or plant part to one ormore bacteria or enzymes. Bacteria such as Rhodococcus spp., Pseudomonaschloroaphis, and Brevibacterium ketoglutamicum may be used. However, theuse of live organisms can be fraught with problems since they aresensitive to environmental conditions, and control of the process (e.g.reversibility) may be difficult. In addition, the public may bereluctant to consume produce that has been “infected” with bacteria.

Glycine betaine is known to be used as an osmolyte regulator whenapplied to fruit before harvesting so as to prevent or reduce blemishesor damage of the skin of the fruit (e.g. to reduce cherry cracking; seeU.S. Pat. No. 5,972,840 to Mottram, the complete contents of which arehereby incorporated by reference). However, U.S. Pat. No. 5,972,840makes no mention of the use of glycine betaine to forestall ripening orprevent pathogen entry of fruit or other produce.

In addition to its role in ripening, ethylene plays a role inabscission, the process of a plant intentionally shedding an organ.Plants naturally use the process of abscission to drop fruit at the endof a growing season. While this process is beneficial in nature,dropping fruit before harvest leads to large losses in the agriculturalworld. U.S. Pat. Nos. 5,869,424 and 6,153,559 addresses this issuethrough the application of aminoethoxyvinylglycine (commercially namedRETAIN®) and its precursor N-acetyl aminoethoxyvinylglycine,respectively, to inhibit ethylene biosynthesis. An alternative strategywas described in US patent US 2012/0101164 A1 where application of1-naphthaleneacetic acid (NAA) also delayed the abscission of fruit.However, these approaches act through different mechanistic propertiescompared to glycine betaine.

SUMMARY OF THE INVENTION

The present invention provides methods and compositions for slowing orcontrolling the timing of the ripening, maturation, senescence,abscission or preventing pathogen entry of produce, for example, fruit,vegetables, ornamental plants, etc. The methods involve the applicationof a photosynthetic pigment or membrane stabilizing agent such asglycine betaine to the produce pre- or post-harvest (or both), therebylengthening the time period during which the produce can be transportedand stored before being made available to consumers. The delay inripening or maturation is advantageously reversible: with the passage oftime, the effects of e.g. glycine betaine decrease, and ripening occurs.The methods are especially useful for treating produce that does notrespond well to the application of 1-MCP, e.g. pears. However, theapplication of the stabilizing agent may be used for any and all producein place of or in conjunction with 1-MCP or other agents, in order tomodulate or control the ripening and/or maturation processes, and totemporarily prevent or slow the onset of senescence.

The practice of the invention will advantageously impact the produce(e.g. fruit and vegetable) industry in at least the following ways:

1) The use of the invention allows for the regulation of ripening sothat the maturation process of produce is controlled and/or delayed e.g.senescence is delayed, and reduced amounts of soluble solids aremaintained;

2) The use of the invention decreases the incidence of fruit drop, ororgan abscission, enabling a longer growing season or delaying harvesttime;

3.) The use of the invention as described herein to reduce incidence ofpreharvest, at harvest, or postharvest pathogen entry from bacteria,fungi, or other pathogens of produce.

4) The use of the invention provides an alternative to 1-MCP and theeffects of glycine betaine, unlike 1-MCP, are reversible;

5) The use of the invention decreases rates of damage during shipment ofe.g. pears and/or other commodities due to the preservation ormaintenance of increased firmness after application;

6) Organic preparations can be formulated, which is not possible with1-MCP in certain cases; and

7) The use of glycine betaine as described herein in combination withe.g. low levels of 1-MCP or other products that interfere withethylene-related or unrelated ripening processes advantageously allowsfor a synergistic slowing of ripening via multiple mechanisms. Suitableratios of glycine betaine to another product in a mixture may range frome.g. about 90:10 to 10:90, including e.g. 50:50; or from about 65:35 to35:65; or from about 75:25 to 25:75, etc;

It is an object of this invention to provide a method of delaying orslowing the ripening, maturation, abscission, or senescence of produce,or preventing pathogen entry into produce. The method comprises thesteps of 1) identifying produce for which delaying or slowing ripening,maturation, abscission, senescence, or pathogen entry is desirable, and2) applying a photosynthetic stabilizing agent to said produce. In someembodiments of the invention, the photosynthetic stabilizing agent isglycine betaine. The produce comprises, for example, fruits, vegetables,and/or ornamental plants. In some cases, the produce is a pear such as ad'Anjou pear. The step of applying may be carried out at one or more ofbefore harvest, at harvest, or after harvest. Alternatively, or inaddition, the step of applying may be carried out during shipping.Application may be by spraying, fogging, dipping, or by infiltration.The step of applying may utilize specific concentrations ofphotosynthetic stabilizing and requires human intervention, e.g. humanintervention in one or more of the processes of ripening, maturation,abscission, and/or senescence of produce, and/or preventing pathogenentry into produce.

The invention also provides a method of delaying or slowing theripening, maturation, abscission or senescence of and preventingpathogen entry into pears. The method comprises the steps of 1)identifying pears for which delaying or slowing ripening, maturation,abscission or senescence and preventing pathogen entry is desirable, and2) applying a photosynthetic stabilizing agent to said pears. In someembodiments, the photosynthetic stabilizing agent is glycine betaine.

The invention further encompasses a composition comprising 1) glycinebetaine, 2) 1-methylcyclopropene (1-MCP), and 3) a carrier suitable forapplication to produce. The invention further encompasses a compositioncomprising 1) glycine betaine, 2), aminoethoxyvinylglycine HCl, and 3) acarrier suitable for application to produce. The invention furtherencompasses a composition comprising 1) glycine betaine, 2),1-naphthaleneacetic acid, and 3) a carrier suitable for application toproduce.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1. Fruit pressure of Bluestim™ (glycine betaine)-treated anduntreated d'Anjou pears. D'Anjou pears were treated on trees withBluestim™ concentrations of 0, 2.5, 5, and 10 lbs/acre on Aug. 17, 2010.Pears were harvested on Sep. 20, 2010 and stored in commercialcontrolled atmosphere (CA) facilities. Pears were removed from storageon Jan. 7, 2011.

FIG. 2. Percent Brix of Bluestim™-treated pears after CA storage.Percent Brix were measured for all pear treatments after they wereremoved from the commercial CA storage conditions.

FIG. 3. Fruit pressure of Bluestim™ (glycine betaine)-treated anduntreated d'Anjou pears. D'Anjou pears were treated on trees withBluestim™ concentrations of 0 and 7.5 lbs/acre before harvest. Pearswere harvested during the fall and stored in commercial controlledatmosphere (CA) facilities. Pears were removed from storageapproximately 3 months later. Fruit firmness and % Brix were measuredapproximately I month after removal from storage.

DETAILED DESCRIPTION

The present invention provides methods and compositions that temporarilyprevents or delays one or more of the ripening, maturation, abscissionand senescence of, and/or prevention of pathogen entry into, producesuch as fruits and vegetables. Further, the methods and compositionspreserve the freshness of ornamental plants. The compositions comprise aphotosynthetic membrane and/or pigment stabilizing agent such as glycinebetaine, a natural product found in most fruit, vegetable and animalspecies. Without being bound by theory, it is believed that the externalapplication of such an agent to produce that is not yet ripe or which isin the process of ripening slows the breakdown of photosyntheticpigment, which usually accompanies and/or participates in the ripeningprocess. As a result of the slowing of photosynthetic membrane and/orpigment and protein complex breakdown, ripening or maturation of theproduce is also slowed. However, the effect is not permanent.

With time, the effects of the application decrease or reverse, andripening/abscission/senescence/maturation proceeds in a normal manner.The application of e.g. glycine betaine thus reversibly prolongs thefirmness of produce and retards the conversion of starches to solublesugars, both of which are considered in the art to be indicators ofripening. The use of such an agent is particularly advantageous for usewith e.g. pears, which can be problematic since they often fail to ripenat all (or only minimally) after exposure to conventional agents such as1-MCP. The prolongation of ripening and/or maturation and/or the delayof abscission and/or the delay of senescence facilitates storage andtransportation of produce, both with respect to time (i.e. the time ofstorage can be extended without loss due to spoilage), and with respectto decreasing damage that otherwise occurs during shipping and handlingof softened, softening or wilting produce. Additionally, stabilizationof membranes promotes the ability of the produce to withstand infectionby pathogens.

By “prevention of pathogen entry” or “prevention of pathogen entry into”we mean that the use of glycine betaine slows or prevents or lessens theamount or extent of pathogen invasion or infestation of the plant orproduce, compared to corresponding plants/plant tissue that is/areexposed to the pathogen under comparable conditions but withouttreatment with glycine betaine. Such invasion or infection may be, forexample, actual entry of the pathogen (e.g. by boring, chewing, enzymesecretion, etc.) into plant tissue; or the eating of plant tissue at ornear the surface; or the insertion of body parts of the pathogen intoplant tissue (e.g. feeding or reproductive appendages); or the placementof reproductive means (e.g. the laying of eggs) in or on plant tissue;or the establishment of a colony in or on the plant or plant tissue,etc. Pathogens, pests and parasites that may be prevented from infectingplants, produce, etc. by the use of glycine betaine include but are notlimited to: various rusts and blights; insects; nemartodes; fungi,bacteria, viruses, mildews, aphids, etc.

In one embodiment, the photosynthetic stabilizing agent is glycinebetaine. While much of the discussion herein refers to glycine betaine,the discussion can apply equally to other photosynthetic stabilizingagents or precursors of glycine betaine such as choline orbetainealdehyde, examples of which include but are not limited to:chlorophyllase inhibitors such as diisopropyl fluorophosphates,thylakoid membrane stabilizers such as spermine and 1,3-diaminopropaneetc.

Glycine betaine (also known as betaine, trimethylglycine or “TMG”,betaine anhydrous, N,N,N-trimethylglycine, etc.) is a natural compoundthat occurs in plants and animals as either glycine betaine or acholine-containing glycine betaine compound which can be enzymaticallycleaved to form glycine betaine. First discovered in sugar beets,glycine betaine is produced abundantly as a byproduct of the manufactureof e.g. molasses and sugar from sugar beets and sugar cane.Significantly, this substance is known to be non-toxic and thus its useon comestible substances is safe with respect to consumption by humansor other animals. Betaine is already known for use as an animal feedsupplement; in products for skin, hair and oral care (e.g. as amoisturizer and protectant); as a de-icing agent; as a sportsperformance enhancing agent that is added to e.g. sports drinks; and forother purposes. It is thus readily available commercially, for example,from Finnfeeds Finland, in Naantali, Finland, as the product BLUESTIM®(97% solution of betaine), and in various forms from the Daniscocompany; or as “Glycine Betaine” from Sunwin in China; or as “Prostim”“GREENSTIM®” or “Jinsaeng” from Agrogenesis in Finland (see, for examplethe websites located atwww.agro-genesis.cam/product_environmentalcare_turf.htmland and thewebsite located at www.agro-genesis.com/product_cropscience_phyto.html;etc.

The glycine betaine that is used as described herein may be “organic”,by which we mean that the source of the glycine betaine can be producedusing methods that do not involve synthetic inputs such as syntheticpesticides, chemical fertilizers, etc. and, optionally, are notgenetically modified or processed using irradiation, industrialsolvents, or chemical food additives. Various standards of “organic”have been established in many countries and locations, including theUnited States, European countries, South American countries, and others.These standards may vary somewhat from location to location, but ingeneral the definition provided above is what is intended when the termis used herein.

With respect to its use on produce as described herein, glycine betainemay be applied “as is” from a commercial source, or may be formulated orreformulated into a preparation that is used for application. Glycinebetaine may be applied, for example, as a liquid solution that contains,e.g. at least about 10, 20, 30, 40, 50, 60, 70, 80, 90, or even about100% glycine betaine, so long as the formulation is suitable forapplication, and so long as the applied solution adheres to or isretained at least partially on the surface of the produce or plant thatis being treated. Further, glycine betaine is not always applied as aliquid; solid forms (e.g. crystals, dusts, aerosols, etc.) of this agentare also contemplated and encompassed by the invention, as are variousgels or other forms. In addition, various substances that are not active(i.e. are inert) with respect to ripening in particular, or to producein general, may be included in such formulations. For example, agentsthat aid in preparation or application of the formulation, or inretention of the glycine betaine on the produce after application, orfor any other reason may be included, e.g. preservatives, colorants,detergents, various emulsifiers, diluents, buffering agents, salts,oils, lipids, waxes, solvents, penetrants, paper wrappings, packaging,storage boxes. Glycine betaine may also be directly infiltrated viavacuum or injected to retard ripening.

In addition, glycine betaine may be used alone or in combination withone or more other active agents which affect/improve/slow, etc. theripening process, or which are beneficial to produce in some other way.By “used together” or “in combination” we mean that the two (or more,i.e. a plurality of) agents may be applied 1) separately one after theother within a relatively short period of time (e.g. simultaneously fromseparate preparations, or within minutes or hours, or the same day, orwithin a few days, etc.); or 2) the two or more agents may be mixedtogether into a single formulation that is applied. Glycine betaine maythus be used in combination with 1-MCP, other anti-ripening agents suchas lysophosphatidylethanolamine; silver agents, aminoethoxyvinylglycine,aminooxyacetic acid, potassium permanganate (ethylene absorbent),aminoethoxyvinylglycine HCl, 1-naphthaleneacetic acid, etc. Additionalactive ingredients or additives that may be present in such compositionsinclude but are not limited to: various antimicrobial substances;various antifungal agents; various insecticides; various pesticides;hormones, biotic applications, etc. For example, see U.S. Pat. Nos.5,110,341; 6,177,072; and 7,150,884, the complete contents of each ofwhich are hereby incorporated by reference, for a description of someexemplary ingredients or additives with which glycine betaine may becombined. Generally, the amount of glycine betaine in such a compositionis from about 1 to about 99%.

Those of skill in the art will recognize that the amount of glycinebetaine that is applied may vary from circumstance to circumstance, e.g.according to the produce that is being treated, according to the desiredamount of delay that is desired, according to the environment duringdelivery (e.g. in an orchard vs in a shipping container), according toweather conditions, etc. In general, the amount for application “in thefield” e.g. in an orchard, filed, etc. is from about 0.5 to about 50, orfrom about 1 to about 15, or from about 2.5 to 10, e.g. about 1, 2, 3,4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18 19, 20, 25, 30, 35,40, 45 or 50 pounds per acre. For other scenarios, such as in shippingcontainers, about 20, 40, 60, 80 or 100 pounds is applied.

According to the invention, glycine betaine may be applied to any typeof produce, (including fruit, vegetables, and ornamental plants) forwhich it is desirable to slow maturation. Thus, the term “produce” asused herein is intended to be interpreted broadly, and to includeplant-based agricultural products that are typically referred to asfruits, vegetables, grasses, grains, ornamental plants, bulbs, nuts,etc.

With respect to fruit, those of skill in the art will recognize that theword “fruit” may have a variety of meanings. For example, standarddefinitions include: 1. edible part of plant: an edible part of a plant,usually fleshy and containing seeds; 2. ovary of a plant: the ripenedseed-bearing ovary of a plant; and 3. produce: the produce of any plantgrown or harvested by humans. In broad terms, “fruit” may refer to astructure of a plant that contains its seeds. In non-technical usage,such as food preparation, “fruit” may refer to the fleshyseed-associated structures of certain plants that are sweet and ediblein the raw state, such as apples, oranges, grapes, strawberries, juniperberries, bananas, etc., although non-sweet or less sweet fleshystructures (e.g. root vegetables such as carrots, beets, sweet potatoes,yams, etc.) are also included. Seed-associated structures that do notfit these informal criteria may be referred to by other names, such asvegetables, pods, ears, etc. In the botany of flowering plants, a“fruit” is a part that derives from specific tissues of the flower,mainly one or more ovaries. Taken strictly, this definition excludesmany structures that are “fruits” in the common sense of the term, whileincluding many structures that are not commonly called “fruits”, such asbean or pea pods, corn kernels, tomatoes, etc. all of which areencompassed by the present invention.

Many fruits that, in a botanical sense, are true fruits are actuallytreated as vegetables in cooking and food preparation, includingcucurbits (e.g., squash, pumpkin, gourds, melons, cucumbers, etc.),tomatoes, peas, beans, corn, eggplant, and sweet pepper, as wells assome spices, such as allspice and chilies.

As used herein, the term “fruit” refers to all plant productsencompassed by the botanical, culinary and common meanings of the word.

Exemplary types of fleshy, simple fruits encompassed by the inventioninclude but are not limited to: stone fruit or drupe (e.g. plum, cherry,peach, apricot, olive, mango, etc.); pome fruits of the family Rosaceae,(including apples, pears, rosehips, saskatoon berry, etc.); aggregatefruits such as achenes (e.g. strawberry), follicles, drupelets(raspberry and blackberry), and various other berries; multiple fruitssuch as pineapple, fig, mulberry, osage-orange, breadfruit, hedge apple,etc; citrus fruits such as oranges, lemons limes, grapefruits, kumquats,tangelos, ugh fruit, tangerines, tangelos, minnolas, etc.; so-called“true” berries such as black current, red current, gooseberry, tomato,eggplant, guava, lucuma, chilis, pomegranates, kiwi fruit, grape,cranberry, blueberry, etc.; including both seeded and seedlessvarieties, as well as hybrid and genetically altered or manipulatedvarieties; and others such a avocados, persimmons, etc.

In one embodiment of the invention, the fruit that is treated using themethods and compositions of the invention is a pear. By “pear” we meanthe fruit of a member of the genes Pyrus, examples of which include butare not limited to: Pyrus amygdaliformis (Almond-leafed pear); Pyrusarmeniacifolia; Pyrus boissieriana; Pyrus bourgaeana (Iberian pear);Pyrus×bretschneideri (Chinese white pear; also classified as asubspecies of Pyrus pyrifolia); Pyrus calleryana (Callery pear); Pyruscommunis (European pear); Pyrus communis subsp. communis (European pearcultivars of which include Beurre d'Anjou, Bartlett and Beurre Bosc);Pyrus communis subsp. caucasica (syn. P. caucasica); Pyrus communissubsp. pyraster (wild European pear (syn. Pyrus pyraster); Pyrus cordata(Plymouth pear); Pyrus cossonii (Algerian pear); Pyrus dimorphophylla;Pyrus elaeagnifolia (oleaster-leafed pear); Pyrus fauriei; Pyrusgharbiana; Pyrus glabra; Pyrus hondoensis; Pyrus koehnei (evergreen pearof southern China and Taiwan); Pyrus korshinskyi; Pyrus mamorensis;Pyrus nivalis (snow pear); Pyrus pashia (Afghan pear); Pyrus×phaeocarpa;Pyrus pseudopashia; Pyrus pyrifolia (Nashi pear, Sha Li); Pyrus regelii;Pyrus salicifolia (willow-leafed pear); Pyrus×serrulata;Pyrus×sinkiangensis (thought to be an interspecific hybrid betweenP.×bretschneideri and Pyrus communis); Pyrus syriaca; Pyrus ussuriensis(Siberian pear) and Pyrus xerophila.

Major types of pears which are marketed and which may be treated by themethods and compositions of the invention include but are not limitedto: Williams' Bon Chréetien (sold in the United States. as Bartlett),Red Bartlett varieties, d'Anjou, Bosc, Cornice, Abate Fatelle, Concorde,and Seckel pears.

Glycine betaine may also be used to slow the development, ripening,maturation, abscission, or onset or progression of senescence of, or toprevent pathogen entry into, vegetables. As used herein, the term“vegetable” refers to an edible plant or part of a plant, and typicallymeans the leaf, stem, or root of a plant but also includes some “fruits”as well (such as squash, see above). The meaning of this word is largelybased on culinary and cultural tradition and all common meanings thatare recognized by those of skill in the art are encompassed herein.Vegetables may be categorized as allium, brassica or cruciferous,composite, gourds, umbelliferous, amaranth, grass, morning glory,nightshade, legumes, root, leafy green, etc. and all such vegetables maybe treated using the methods and compositions of the invention.

Glycine betaine may also be used to slow the development, maturation,abscission, or senescence of, or prevent pathogen entry into, ornamentalplants and hence to preserve their freshness. As used herein, the phrase“ornamental plant” refers to plants that are grown for decorativepurposes such as e.g., house plants, those grown for cut flowers, etc.as well as certain grasses, annuals, perennials, shrubs, trees, etc. Asused herein the phrase has the same meaning that it is generally used inthe horticultural trades.

The application of glycine betaine to produce or plants may be carriedout at any time at which it is desired to delay the onset and/or theprogression of development, ripening, maturation, abscission, pathogenentry etc. In addition, various life stages of plants may be treated,e.g. sprouts, shoots or seedlings, etc. The methods of the inventioninvolve identifying produce for which a delay in ripening, maturation,abscission, senescence, or pathogen entry is desired or suitable, andtreating the produce as described herein. Accordingly, when treatingproduce, the produce may still be on the tree (or other plant from whichit develops and grows), or may have already been harvested. The producemay be in any “stage” of ripening or maturation. For example, theproduce (e.g. after harvest in the orchard, after harvest and whenpacked in a shipping container, etc.) may have characteristics ofimmature, “green” or “un-ripe” produce such as very low sugar content,high stringency, high “firmness”, lack of colorful pigment, undevelopedor developing buds that have not bloomed, etc. Alternatively, theproduce may have begun to mature or ripen in that e.g. the sugar contenthas begun to increase, firmness has begun to dissipate, the size hasincreased, blooming or flowering has (at least partially) commenced,etc. but the process is not complete. In yet other embodiments, theproduce may be approaching maturity, but the application of glycinebetaine causes suspension of further progression of sugar increase,softening, etc. so that the produce is “arrested” in a ripened or maturestate and spoilage and senescence is prevented or slowed or deferred. Inshort, glycine betaine may be applied to produce at a stage prior to theonset of degradation of photosynthetic pigments in order to delayfurther ripening or maturation of the produce. In addition, glycinebetaine may be applied repeatedly to produce or plants in order toextend its effects over a longer period of time.

In one embodiment, the methods of the invention are carried out before afruit is “ripe”, i.e. before the fruit has a soluble solids content or“SSC” (which generally corresponds to sugar content) that is generallyrecognized in the art as suitable for a fruit of interest. Those ofskill in the art will recognize that this number or range varies widelyfrom fruit to fruit. Percent sugar content is typically measured, forexample, in degrees Brix (symbol ° Bx) which is the sugar content of anaqueous solution. One degree Brix is 1 gram of sucrose in 100 grams ofsolution and represents the strength of the solution as percentage byweight (% w/w) (strictly speaking, by mass). If the solution containsdissolved solids other than pure sucrose, then the ° Bx is onlyapproximate the dissolved solid content. Those of skill in the art willrecognize that typical Brix values for fruit and vegetables to whichglycine betaine is applied as described herein will vary widelydepending on the type of fruit or vegetable, and will be able to readilyable to determine suitable values.

SSC values may be determined by various means, for example, byrefractometry, by using various visible/near infrared (Vis/NIR)transmittance techniques, etc.

In addition, the degree of firmness and/or lack thereof may be used asan indicator of ripeness. Fruit thinness is generally measured askg/cm⁻² and suitable ranges of firmness values for unripened and ripefruit and vegetables are well-known to those of skill in the art.Firmness may be measured using a variety of instruments such as variousfruit firmness testers, force gauges, etc.

In addition, other criteria may also be employed to determine the extentof ripeness (or lack thereof) of a fruit, for example, color, aroma,free acids, starch content, internal ethylene, etc.

Those of skill in the art will recognize that all pieces of produce in abatch that is being analyzed for possible treatment according to themethods of the invention may not be at the same stage of ripening at thesame time. Thus, when a term such a “un-ripened fruit” is used, it maybe used in the collective sense in that, for example, most or asignificant portion of the fruit in a batch of interest is not ripe,e.g. at least about 10, 20, 30, 40, 50, 60, 70, 80, 90, or possibly 100%of the fruit meets the criteria set forth herein as “not-ripened” or“partially” or “fully” ripened, etc. as the case may be. In addition,those of skill in the art will recognize that the criteria fordetermining the stage of ripeness or maturation may differ from producetype to produce type, or depending on the use or intended handling ofthe produce type. For example, some apples are considered “ripe” or“mature” or “ready to eat” or “ready to use” e.g. for cooking (at leastby consumers) when they are still very firm (e.g. crisp) and tart, whileother apples, or other fruits, are much sweeter and/or softer when theyare “ready”.

With respect to vegetables and ornamental plants, the same or similarprocedures apply, i.e. glycine betaine may be applied at any stage ofdevelopment at which it is desired to arrest further maturation and/orprevent senescence or spoilage of a plant or plant product. The criteriafor maturation or “readiness” for vegetables and ornamental plants areknown to those of skill in the art and may be used to determine suitabletimes or stages for application. For example, application may bepre-harvest when a plant is still in the field, or post-harvest duringstorage or shipment. Glycine betaine may be applied to leaves and/or toany other accessible part of a plant, e.g. to portions of a plant suchas leaves, pea pods, ears of corn still in the husk, etc., or,post-harvest to roots, peas that have been removed from pods, corn thathas been husked, etc., to name only a few illustrative examples.

In some embodiments, glycine betaine is applied, usually by commercialgrowers, as a liquid solution using methods that are known to those ofskill in the art, e.g. using known spray apparatuses such as spray guns,various pressurized apparatuses, etc._; or may be applied as a “fog” oraerosol e.g. within a storage or shipping compartment; or by dipping orwashing of the produce; or by any other suitable means. However, it isnot necessary that liquid formulations be used. Glycine betaine may alsobe applied as granules or as a “dust” using application technologiesthat are known in the art. Such applications may or may not be timed tocoincide with other conditions such a weather, e.g. application ofgranules or dusts may be carried out soon after a rain or afterapplication of another liquid (e.g. a liquid pesticide) or duringperiods of high humidity, etc., so that the granules/dust will readilyadhere to the fruit (and liquefy, at least to some extent) on theexterior of the fruit.

The application of glycine betaine may be carried out only once, or maybe repeated as needed or desired to maintain a desired status of theproduce. For example, an initial application may be followed bysubsequent applications at intervals of several days, or weeks, ormonths, etc. as required to keep the produce in the desired state. Inaddition, the application of glycine betaine may be preceded, followedor alternated with the application of other agents such as otheranti-ripening, or anti-maturation agents; or ripening or maturationagents may be applied to hasten the reversal of the effects of glycinebetaine. In general, the effects of a single application of glycine havea duration as long as from about 5 to about 30 days, and may be as shortas from about 1 to about 5 days.

In another embodiment of the invention, glycine betaine is applied toproduce or crops prior to harvest while they are “in the field”(orchard, etc.) in order to protect the produce or crops fromenvironmental stress such as drought, excessive heat, excessive cold,sun damage, etc. The application under these circumstances isessentially the same as described above, but the produce or crop isidentified as being at risk of damage due to untoward environmentalconditions such a weather, e.g. lack of rain, high temperatures, etc.

In another embodiment of the invention, glycine betaine is applied toproduce or crops prior to harvest while they are “in the field”(orchard, etc.) in order to delay the abscission or fruit drop of theproduce. This application may occur one or multiple times in a growingseason.

In some embodiments of the invention, glycine betaine preparations areprepared essentially for use by commercial growers and/or entities whichare involved in the storage or shipping of produce. However, in otherembodiments, the formulations are marketed and sold directly toconsumers, e.g. to a consumer who wishes to delay the ripening orspoilage of fruit. Due to the known safety of glycine betaine, provisionof such preparations directly to consumers does not pose a health risk,even if, for example, directions supplied with the preparation are notfollowed precisely, or if a solution of glycine betaine is spilled,consumed by a pet, etc. For example, small-scale, sprayable liquidsolutions may be provided, and/or concentrated solutions or granuleswhich can be diluted or dissolved and applied by spraying or by othermeans (e.g. as a fruit “wash”) may be provided. This option may beparticularly appealing to consumers who like to buy produce ahead oftime (e.g. to take advantage of a sale, to buy produce during peakseason, etc.), and store it for a period of e.g. weeks or months, or whogrow and harvest their own fruits, vegetables, flowers, etc.

In addition, “green” and/or “organic” and/or “environmentally sensitive”products comprising glycine betaine may be developed and marketed foruse, for example, by growers, suppliers, consumers, etc., especiallysince glycine betaine is a natural by-product of other processes andthus its use can truly be deemed efficient, not wasteful, etc.

EXAMPLES Example 1

Research described in this Example shows that application of a naturalbyproduct of molasses, glycine betaine, to d'Anjou pears before harvestyields firmer pears compared to untreated pears stored in the samepost-harvest conditions (FIGS. 1 and 3). Application increased the fruitfirmness in a dose dependent manner, with applications of 10 lbs/acreyielding the firmest pears.

Glycine betaine treated pears also display a decrease in soluble sugarsmeasured in % BRIX in a dose dependent manner (FIGS. 2 and 3). Bothfirmness and % BRIX are used as an indicator of fruit ripeness.

As time passes, however, glycine betaine treated pears soften andexhibit increases in % BRIX, indicating that the effect is temporary,and that ripening eventually occurs after treatment.

While the invention has been described in terms of its preferredembodiments, those skilled in the art will recognize that the inventioncan be practiced with modification within the spirit and scope of theappended claims. Accordingly, the present invention should not belimited to the embodiments as described above, but should furtherinclude all modifications and equivalents thereof within the spirit andscope of the description provided herein.

1-12. (canceled)
 13. A method of delaying or slowing the ripening ormaturation of apples, comprising the steps of identifying apples forwhich delaying or slowing ripening or maturation, senescence, orpathogen entry is desirable; and applying glycine betaine to saidapples, wherein said step of applying is carried out at or afterharvest. 14-15. (canceled)
 16. The method of claim 13 wherein saidapplying step includes applying an anti-ripening agent selected from thegroup consisting of 1-methylcyclopropene (1-MCP) andaminoethoxyvinylglycine HCl.